Integrated Copy Number and Expression Analysis Identifies Profiles of Whole-Arm Chromosomal Alterations and Subgroups with Favorable Outcome in Ovarian Clear Cell Carcinomas

Ovarian clear cell carcinoma (CCC) is generally associated with chemoresistance and poor clinical outcome, even with early diagnosis; whereas high-grade serous carcinomas (SCs) and endometrioid carcinomas (ECs) are commonly chemosensitive at advanced stages. Although an integrated genomic analysis of SC has been performed, conclusive views on copy number and expression profiles for CCC are still limited. In this study, we performed single nucleotide polymorphism analysis with 57 epithelial ovarian cancers (31 CCCs, 14 SCs, and 12 ECs) and microarray expression analysis with 55 cancers (25 CCCs, 16 SCs, and 14 ECs). We then evaluated PIK3CA mutations and ARID1A expression in CCCs. SNP array analysis classified 13% of CCCs into a cluster with high frequency and focal range of copy number alterations (CNAs), significantly lower than for SCs (93%, P < 0.01) and ECs (50%, P = 0.017). The ratio of whole-arm to all CNAs was higher in CCCs (46.9%) than SCs (21.7%; P < 0.0001). SCs with loss of heterozygosity (LOH) of BRCA1 (85%) also had LOH of NF1 and TP53, and LOH of BRCA2 (62%) coexisted with LOH of RB1 and TP53. Microarray analysis classified CCCs into three clusters. One cluster (CCC-2, n = 10) showed more favorable prognosis than the CCC-1 and CCC-3 clusters (P = 0.041). Coexistent alterations of PIK3CA and ARID1A were more common in CCC-1 and CCC-3 (7/11, 64%) than in CCC-2 (0/10, 0%; P < 0.01). Being in cluster CCC-2 was an independent favorable prognostic factor in CCC. In conclusion, CCC was characterized by a high ratio of whole-arm CNAs; whereas CNAs in SC were mainly focal, but preferentially caused LOH of well-known tumor suppressor genes. As such, expression profiles might be useful for sub-classification of CCC, and might provide useful information on prognosis.     

Human α‐amino‐β‐carboxymuconate‐ε‐semialdehyde decarboxylase (ACMSD): A structural and mechanistic unveiling

Human α‐amino‐β‐carboxymuconate‐ε‐semialdehyde decarboxylase determines the fate of tryptophan metabolites in the kynurenine pathway by controlling the quinolinate levels for de novo nicotinamide adenine dinucleotide biosynthesis. The unstable nature of its substrate has made gaining insight into its reaction mechanism difficult. Our electron paramagnetic resonance (EPR) spectroscopic study on the Cu‐substituted human enzyme suggests that the native substrate does not directly ligate to the metal ion. Substrate binding did not result in a change of either the hyperfine structure or the super‐hyperfine structure of the EPR spectrum. We also determined the crystal structure of the human enzyme in its native catalytically active state (at 1.99 Å resolution), a substrate analogue‐bound form (2.50 Å resolution), and a selected active site mutant form with one of the putative substrate binding residues altered (2.32 Å resolution). These structures illustrate that each asymmetric unit contains three pairs of dimers. Consistent with the EPR findings, the ligand‐bound complex structure shows that the substrate analogue does not directly coordinate to the metal ion but is bound to the active site by two arginine residues through noncovalent interactions. Proteins 2015; 83:178–187. © 2014 Wiley Periodicals, Inc.     

Possibility of Production of Amino Acids by Impact Reaction Using a Light-Gas Gun as a Simulation of Asteroid Impacts

In order to investigate impact production of carbonaceous products by asteroids on Titan and other satellites and planets, simulation experiments were carried out using a 2-stage light gas gun. A small polycarbonate or metal bullet with about 6.5 km/s was injected into a pressurized target chamber filled with 1 atm of nitrogen gas, to collide with a ice + iron target or an iron target or a ice + hexane + iron target. After the impact, black soot including fine particles was deposited on the chamber wall. The soot was carefully collected and analyzed by High Performance Liquid Chromatography (HPLC), Fourier Transform Infrared Spectroscopy (FT-IR), and Laser Desorption Time-of-Flight Mass Spectrometry (LD-ToF-MS). As a result of the HPLC analysis, about 0.04–8 pmol of glycine, and a lesser amount of alanine were found in the samples when the ice + hexane + iron target was used. In case of the ice + iron target and the iron target, less amino acids were produced. The identification of the amino acids was also supported by FTIR and LD-ToF-MS analysis.     

Extensive Chromosome Homoeology among Brassiceae Species Were Revealed by Comparative Genetic Mapping with High-Density EST-Based SNP Markers in Radish (Raphanus sativus L.)

A linkage map of expressed sequence tag (EST)-based markers in radish (Raphanus sativus L.) was constructed using a low-cost and high-efficiency single-nucleotide polymorphism (SNP) genotyping method named multiplex polymerase chain reaction–mixed probe dot-blot analysis developed in this study. Seven hundred and forty-six SNP markers derived from EST sequences of R. sativus were assigned to nine linkage groups with a total length of 806.7 cM. By BLASTN, 726 markers were found to have homologous genes in Arabidopsis thaliana, and 72 syntenic regions, which have great potential for utilizing genomic information of the model species A. thaliana in basic and applied genetics of R. sativus, were identified. By construction and analysis of the genome structures of R. sativus based on the 24 genomic blocks within the Brassicaceae ancestral karyotype, 23 of the 24 genomic blocks were detected in the genome of R. sativus, and half of them were found to be triplicated. Comparison of the genome structure of R. sativus with those of the A, B, and C genomes of Brassica species and that of Sinapis alba L. revealed extensive chromosome homoeology among Brassiceae species, which would facilitate transfer of the genomic information from one Brassiceae species to another.     

A Novel Antibody Engineering Strategy for Making Monovalent Bispecific Heterodimeric IgG Antibodies by Electrostatic Steering Mechanism

Producing pure and well behaved bispecific antibodies (bsAbs) on a large scale for preclinical and clinical testing is a challenging task. Here, we describe a new strategy for making monovalent bispecific heterodimeric IgG antibodies in mammalian cells. We applied an electrostatic steering mechanism to engineer antibody light chain-heavy chain (LC-HC) interface residues in such a way that each LC strongly favors its cognate HC when two different HCs and two different LCs are co-expressed in the same cell to assemble a functional bispecific antibody. We produced heterodimeric IgGs from transiently and stably transfected mammalian cells. The engineered heterodimeric IgG molecules maintain the overall IgG structure with correct LC-HC pairings, bind to two different antigens with comparable affinity when compared with their parental antibodies, and retain the functionality of parental antibodies in biological assays. In addition, the bispecific heterodimeric IgG derived from anti-HER2 and anti-EGF receptor (EGFR) antibody was shown to induce a higher level of receptor internalization than the combination of two parental antibodies. Mouse xenograft BxPC-3, Panc-1, and Calu-3 human tumor models showed that the heterodimeric IgGs strongly inhibited tumor growth. The described approach can be used to generate tools from two pre-existent antibodies and explore the potential of bispecific antibodies. The asymmetrically engineered Fc variants for antibody-dependent cellular cytotoxicity enhancement could be embedded in monovalent bispecific heterodimeric IgG to make best-in-class therapeutic antibodies.     

On the reflex coactivation of ankle flexor and extensor muscles induced by a sudden drop of support surface during walking in humans.

Recent studies have revealed that the stretch reflex responses of both ankle flexor and extensor muscles are coaugmented in the early stance phase of human walking, suggesting that these coaugmented reflex responses contribute to secure foot stabilization around the heel strike. To test whether the reflex responses mediated by the stretch reflex pathway are actually induced in both the ankle flexor and extensor muscles when the supportive surface is suddenly destabilized, we investigated the electromyographic (EMG) responses induced after a sudden drop of the supportive surface at the early stance phase of human walking. While subjects walked on a walkway, the specially designed movable supportive surface was unexpectedly dropped 10 mm during the early stance phase. The results showed that short-latency reflex EMG responses after the impact of the drop (<50 ms) were consistently observed in both the ankle flexor and extensor muscles in the perturbed leg. Of particular interest was that a distinct response appeared in the tibialis anterior muscle, although this muscle showed little background EMG activity during the stance phase. These results indicated that the reflex activities in the ankle muscles certainly acted when the supportive surface was unexpectedly destabilized just after the heel strike during walking. These reflex responses were most probably mediated by the facilitated stretch reflex pathways of the ankle muscles at the early stance phase and were suggested to be relevant to secure stabilization around the ankle joint during human walking.     

Age-related changes of the stretch reflex excitability in human ankle muscles

The purpose of this study was to characterize the effects of aging on the stretch reflex in the ankle muscles, and in particular to compare the effects on the ankle dorsi-flexor (tibialis anterior: TA) and the plantar-flexor (soleus: SOL). Stretch reflex responses were elicited in the TA and SOL at rest and during weak voluntary contractions in 20 elderly and 23 young volunteers. The results indicated that, in the TA muscle, the elderly group had a remarkably larger long-latency reflex (LLR), whereas no aging effect was found in the short latency reflex (SLR). These results were very different from those in the SOL muscle, which showed significant aging effects in the SLR and medium latency reflex (MLR), but not in the LLR. Given the fact that the LLR of the TA stretch reflex includes the cortical pathway, it is probable that the effects of aging on the TA stretch reflex involve alterations not only at the spinal level but also at the cortical level. The present results indicate that the stretch reflexes of each of the ankle antagonistic muscles are affected differently by aging, which might have relevance to the neural properties of each muscle.     

Gastrointestinal parasites of the chimpanzee population introduced onto Rubondo Island National Park,Tanzania

The release of any species into a novel environment can evoke transmission of parasites that do not normally parasitize the host as well as potentially introducing new parasites into the environment. Species introductions potentially incur such risks, yet little is currently known about the parasite fauna of introduced primate species over the long term. We describe the results of long‐term monitoring of the intestinal parasite fauna of an unprovisioned, reproducing population of chimpanzees introduced 40 years earlier (1966–1969) onto Rubondo Island in Lake Victoria, Tanzania, a non‐native habitat for chimpanzees. Two parasitological surveys (March 1997–October 1998 and October 2002–December 2005) identified Entamoeba spp. including E. coli, Iodamoeba buetschlii, Troglodytella abrassarti, Chilomastix mesnili, Trichuris sp., Anatrichosoma sp., Strongyloides spp., Strongylida fam. gen. sp., Enterobius anthropopitheci, Subulura sp., Ascarididae gen. sp., and Protospirura muricola. The parasite fauna of the Rubondo chimpanzees is similar to wild chimpanzees living in their natural habitats, but Rubondo chimpanzees have a lower prevalence of strongylids (9%, 3.8%) and a higher prevalence of E. anthropopitheci (8.6%, 17.9%) than reported elsewhere. Species prevalence was similar between our two surveys, with the exception of Strongyloides spp. being higher in the first survey. None of these species are considered to pose a serious health risk to chimpanzees, but continued monitoring of the population and surveys of the parasitic fauna of the two coinhabitant primate species and other animals, natural reservoir hosts of some of the same parasites, is important to better understand the dynamics of host–parasite ecology and potential long‐term implications for chimpanzees introduced into a new habitat. Am. J. Primatol. 72:307–316, 2010. © 2009 Wiley‐Liss, Inc.